Exploring novel synthesis strategies for magnetic beads to extract nucleic acids is of great significance in the field of in vitro diagnostics. In the present research, monodisperse magnetic mesoporous silica beads were synthesized via the thermolysis reaction of Fe(acac)3 by using large-pore dendritic silica colloids as templates, and were further functionalized with a highly pH-sensitive histidine-glutamate co-oligopeptide for deoxyribonucleic acid extraction. The large-pore dendritic silica colloid scaffolds were utilized for high-density incorporation of superparamagnetic iron oxide nanoparticles within the vertical channels. The morphology and properties of the as-prepared pH-sensitive oligopeptide magnetic mesoporous silica beads were evaluated by transmission electron microscopy, scanning electron microscopy, vibrating sample magnetometry, X-ray photoelectron spectroscopy, X-ray diffraction testing and so on. The average size of the obtained magnetic beads was 370 nm in diameter with a narrow size distribution. The saturation magnetization and magnetic content of the resultant magnetic beads were 25 emu g-1 and 59%, respectively. Moreover, the magnetic mesoporous silica beads exhibited an obvious pH-responsive behavior. Due to these remarkable features, successful deoxyribonucleic acid capture using the as-prepared pH-sensitive oligopeptide magnetic mesoporous silica beads was achieved.

BACKGROUND:Circulating tumor DNA (ctDNA) is a promising biomarker for monitoring minimal residual disease (MRD), assessing disease status, and guiding treatment in diffuse large B-cell lymphoma (DLBCL). Current ctDNA assays rarely detect immunoglobulin (IG) fusions. This study evaluates a novel assay that simultaneously detects mutations, IG fusions, and IG V(D)J clonality in plasma and cerebrospinal fluid (CSF) to enhance molecular profiling and CNS monitoring. METHODS:A prospective analysis was conducted in 57 DLBCL patients. Genomic alterations in plasma and CSF ctDNA were compared to those in tumor tissue using targeted next-generation sequencing (NGS). RESULTS:Mutations, IG fusions, and IG V(D)J clonality were detected in 100%, 72.2%, and 78.6% of plasma ctDNA samples, respectively. Plasma ctDNA also revealed additional mutations absent in tumor tissue, reflecting clonal heterogeneity. The incorporation of IG fusion detection enabled molecular subtyping without requiring FISH. In CSF, ctDNA analysis identified genomic alterations in 8 cases, whereas conventional imaging and cytology confirmed CNS involvement in only 3, demonstrating the superior sensitivity of ctDNA for CNS surveillance. CONCLUSION:This ctDNA assay offers a non-invasive, integrated approach for genomic profiling and disease monitoring in DLBCL, with improved sensitivity for CNS detection and potential to inform personalized treatment strategies.

T-cell receptor (TCR) loci undergo complex V(D)J rearrangements during T-cell maturation. However, errors in this process can cause TCR promoters and enhancers to aberrantly fuse with oncogenes, leading to their overexpression and the development of T-cell acute lymphoblastic leukemia (T-ALL). Unexpectedly, in a retrospective study of 97 pediatric B-ALL cases, all diagnosed according to WHO criteria at a single institution, we identified a subgroup of B-ALL harboring TCR fusion. These fusions involved diverse partner genes, including LINC01656, TBC1D10B, TCL1A/TCL1B, CDKN2A, CEBPB/G, and AHI1. Patients with TCR fusions showed a gene expression profile enriched in hematopoietic stem cell (HSC) signatures and frequently carried other adverse genomic markers, such as CDKN2A/B alterations and RAS pathway activation. Clinically, TCR-rearranged B-ALL patients were associated with increased risk (P = 0.005) and had higher minimal residual disease (MRD) at day 19 (P = 0.034), and may benefit from MRD-based risk-directed therapy. Our study highlights frequent TCR fusions in B-ALL, suggesting a stem cell-derived leukemia with potential differentiation into both B-cell and T-cell lineages.

Introduction B-cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. Despite the significant advances in risk classification and improved treatment, recurrence still occurs in 15%-20% of cases. Therefore, identifying new biomarkers capable of predicting the risk of BCP-ALL is crucial for improving outcomes. Rearrangements of the T-cell receptor alpha-delta locus (TRA/D) and immunoglobulin (IG) loci are well-established diagnostic markers for T-cell acute lymphoblastic leukemia (T-ALL) and B-cell acute lymphoblastic leukemia (B-ALL), respectively. Chromosomal translocations involving the TRA/D at the 14q11.2 are found in around 17% of T-ALL but are rare in B-ALL. Herein, we use next-generation sequencing to detect TRA/D rearrangements and describe the clinical and biological characteristics of patients with B-cell ALL and TRA/D rearrangements. Methods We conducted a retrospective study in a single institution involving 97 pediatric B-ALL patients diagnosed between June 2023 and January 2024 to identify TRA/D rearrangements using a targeted DNA next-generation sequencing (NGS) which highly sensitive to detect translocations at the genomic level and use this approach emploings probes spanning the entire IG/TR gene regions. Bone marrow aspirate, immunophenotyping, G-banding karyotype, fluorescent in situ hybridization (FISH), whole-transcriptome RNA sequencing (RNA-seq) and whole-genome sequencing (WGS) were performed for all 97 patients. The involvement of the TRA/D locus was confirmed by FISH and Sanger sequencing. The relative partner gene expression levels (fragments per kilobase of transcript per million mapped reads, FPKM) were estimated based on supporting reads retrieved from the transcriptome RNA-seq datasets and were compared to those with B-ALL without TRA/D rearrangements. We further characterized the expression of hematopoietic stem cells (HSCs)-related genes in this type of TRA/D rearrangements by gene set enrichment analysis (GSEA). Results Surprisingly, we revealed that 6/97 (6.18%) cases of B-cell ALL harbored TRA/D rearrangements. These rearrangements were associated with various partner genes including TCL1A/B, CEBPA, and CDKN2A. Patients with TRA/D rearrangements exhibited a gene expression profile enriched in hematopoietic stem cell features and, in some cases, had additional activating mutations in RAS signaling pathway genes. These tumors often expressed mixed lineage antigens. Compared with patients without TRA/D rearrangements, patients with TRA/D rearrangement were significantly more likely to be classified as intermediate risk (P = 0.047), havinghigher frequency of ZNF384/362 rearrangements (P < 0.001), and s measurable residual disease (MRD) levels above 1% at day 19 (P = 0.034). Conclusions Our observations indicate that TRA/D rearranged may represent a distinct subgroup of pediatric B-ALL, characterized by unique gene expression profiles, immunophenotypes, and prognostic features which have prognostic implications for patients classified as intermediate risk.

Diffuse large B cell lymphoma (DLBCL) expresses abundant programmed death ligand 1 (PD-L1), which shields tumor cells from immune attacks through the PD-L1/PD-1 signaling axis. The mechanism of PD-L1 overexpression includes the deletion of the 3'end of PD-L1, which increases its mRNA stability, and the gain or amplification of PD-L1. Previous studies found two cases of DLBCL carrying an IGH::PD-L1 by whole genome sequencing. We describe two more such cases by a targeted DNA next-generation sequencing (NGS) capable of detecting IGH rearrangements, leading to PD-L1 overexpression. DLBCL with PD-L1 overexpression is often resistant to R-CHOP (rituximab, cyclophosphamide, doxorubicin hydrochloride, vincristine and prednisolone). Our patients responded to a combination of R-CHOP and a PD-1 inhibitor.

The synuclein family, consisting of α-, β-, and γ-synuclein, is primarily expressed in neurons. Mutations of α- and β-synuclein have been linked to Parkinson’s disease and dementia with Lewy bodies, respectively. Recent studies have shown that synucleins are upregulated in various tumors, including breast, ovarian, meningioma, and melanoma, and high synuclein expression is associated with poor prognosis and drug resistance. We report a novel rearrangement of β-synuclein in a pediatric T-cell acute lymphoblastic leukemia (T-ALL) case, where β-synuclein (SNCB) is fused in-frame with ETS variant transcription factor 6 (ETV6), a gene frequently rearranged in acute leukemia including acute myeloid leukemia (AML), B-cell acute lymphoblastic leukemia (B-ALL), and T-ALL. An additional case of β-synuclein rearrangement was identified in a squamous cell carcinoma of the lung through analysis of the public TCGA database. Both rearrangements involve the C-terminal of β-synuclein. Since β-synuclein shares extensive amino acid similarities with α-synuclein and α-synuclein binds to 14-3-3, an important regulator of apoptosis, the rearranged β-synuclein may contribute to tumorigenesis by deregulating apoptosis. In addition, overexpression of synucleins has been shown to increase cell proliferation, suggesting that the rearranged β-synuclein may also deregulate the cell cycle.

Abstract Purpose The synuclein family includes a-, b-, g-synuclein and is predominantly expressed in neurons. a- and b-synuclein is mutated in Parkinson's disease and dementia with Lewy bodies. Recent studies found up-regulation of the synucleins in several tumors including breast cancer, ovarian cancer, meningioma, and melanoma, and the high level of synuclein was associated with poor prognosis and drug resistance. We describe here a novel intragenic rearrangement of b-synuclein in pediatric T-cell acute lymphoblastic leukemia (T-ALL). An additional case of b-synuclein rearrangement was found in a squamous cell carcinoma of the lung by searching the public TCGA database. Methods Morphological evaluation and immunohistochemistry were used for diagnostic purposes. Karyotype analysis, targeted RNA NGS, FISH, and RT-PCR were used to identify the fusion transcript. Results A pediatric T-ALL carried a translocation of chromosomes 5 and 12, resulting in an in-frame fusion between the b-synuclein (SNCB) and the ETS variant transcription factor 6 (ETV6), a gene frequently rearranged in acute myeloid leukemia (AML), B-ALL, and T-ALL. Another SNCB rearrangement involved low-density lipoprotein receptor class A domain containing 3 (LDLRAD3) in a lung carcinoma. Both fusions retained the c-terminal of b-synuclein, a region important for protein interaction. Conclusion We describe the first cases of b-synuclein rearrangement in tumors. Since b-synuclein shares extensive similarity in amino acid sequences with a-synuclein and the a-synuclein binds to 14-3-3, an important regulator of apoptosis, we suspect that the rearranged b-synuclein likely contributes to tumorigenesis by deregulating apoptosis. In addition, the rearranged b-synuclein could deregulate the cell cycle, because overexpression of b-synuclein leads to increased cell proliferation.